Sarcoidosis is a systemic inflammatory disease of unknown etiology and has no cure. It is thought to be caused by an abnormal immune response to either a self or non-self antigen. Sarcoidosis is characterized by a Th1 immune response with interferon-gamma being the predominant cytokine. Despite this knowledge, little is known about the regulators of this aberrant immune response, and current treatment approaches (e.g. systemic corticosteroids and methotrexate) are non-specific and have significant toxicities. Natural killer T (NKT) cells are immunoregulatory T cells that can either promote or suppress immune responses depending on local stimuli. In mice, NKT cells have been found to be protective in several autoimmune diseases. In humans, two independent clinical studies found that the numbers of peripheral blood and lung NKT cells were significantly lower in patients with pulmonary sarcoidosis compared to controls. Thus, NKT cells are abnormally regulated in sarcoidosis. However, large gaps in knowledge exist about the cellular subsets and effector function of NKT cells in this disease. Specific Aims. Because a deficiency of the CD4+ subset of NKT cells may skew immune responses towards Th1 responses (a characteristic of sarcoidosis), we hypothesize that the overall NKT cell deficiency in sarcoidosis is due to a selective loss of the CD4+ NKT cell subset leading to a paucity of Th2 cytokines and a skewing of the cytokine balance in favor of Th1 cytokine-producing CD4- NKT cells. Therefore, we will 1) determine the distribution of CD4+ vs. CD4- NKT cells in sarcoidosis and their cytokine responses after stimulation with CD1d ligands in a cross-sectional study and 2) determine how NKT cell effector function relates to disease severity, progression and phenotype in sarcoidosis subjects and the effect of immune- suppression on NKT cell subsets and cytokine responses in a longitudinal study. Experimental Approach: We will collect peripheral blood mononuclear cells from normal volunteers and subjects with sarcoidosis. Directly ex-vivo we will determine the number of CD4+ and CD4- NKT cells and their cytokine responses to several NKT cell ligands using novel in vitro stimulation methods and multi-color flow cytometric analysis. These responses will be compared to those from matched healthy volunteers. We will also analyze NKT cell subsets and cytokine responses within sarcoidosis subjects longitudinally at ~6 month intervals and correlate these measurements to disease severity. Additionally, we will determine the effect of systemic immunosuppressants on NKT cell subsets and cytokine responses. Finally, we will compare blood and lung NKT cell effector function in a pilot study of 5-10 pulmonary sarcoidosis subjects. Significance of the results. Since NKT cells may play a role in the pathogenesis of sarcoidosis, findings from this study could identify a new cell type that may be targeted in novel treatment strategies using CD1d ligands, which was identified as a major priority area of research in sarcoidosis by a recent NHLBI workshop. PUBLIC HEALTH RELEVANCE: The scientific aims of the clinical project for which we are seeking support from the NIH is to comprehensively analyze the function of a recently identified immunoregulatory cell, the natural killer T cell, in patients with sarcoidosis. One of the overriding goals of this project is to advance the knowledge of the immunology of sarcoidosis so that this may be translated into more specific therapies to reduce suffering of patients and thereby promote lung health and improve patients'lives. In addition, by achieving a better understanding of the immunology of sarcoidosis, we may be able to prevent this lung disease.